The human body has numerous tubular structures that perform necessary and vital physiological functions. Smooth muscle cells are the main cell type responsible for proper physiological function of such tubular structures. The body's hollow tubular structures transport body fluids including, for example, blood, urine, hormones and nutrients. Typically, the tubular structure's role is to actively transport these fluids in a unidirectional way without retrograde movement.
For example, such tubular structures include blood vessels that move blood throughout the body. Blood vessels come in different sizes to propel blood through the body and actively contribute to the blood circulation initiated by the heart. The blood vessels fall in two categories: arteries, which transport oxygenated blood away from the heart, and veins, which return oxygen-depleted blood back to the heart.
The arteries constitute an integral and important part in the circulation of blood. Veins on the other hand are more passive, and are conduits for continuous flow of blood. However, veins have developed special characteristics to minimize backflow, especially in lower extremities in order to fight gravity.
Several attempts have been made to manufacture blood vessels. Most of the attempts use mesenchymal cells either micro spun or printed or simply seeded on a scaffold. They often fail to generate any propulsive force, and function only as conduits -- with some risk of blockage. Such artificial blood vessels may be suitable for vein replacement but have not been shown to be suitable to artery replacement.
Other examples of tubular structures can be found throughout the digestive system. The hollow organs of the digestive system are highly organized and perform coordinated functions to move food and nutrients. To take full advantage of the nutritional values of the different foods ingested, the food has to be at “the right time in the right place” along the digestive tract for proper breakdown, trituration, digestion and absorption. Any slowdown in this propulsive, peristaltic movement, such as localized or segmental paralysis (referred to as “paresis”), can result in localized or segmental inertia that is detrimental for the proper nutrition and can lead to obstruction, infection or morbidity. Neuromuscular diseases of the gastrointestinal track often exhibit this lack of coordinated propulsive movements. Such disorders can manifest themselves in a variety of locations along the digestive track, e.g., at the level of the esophagus, small intestine or large intestine.
The esophageal conduit extends from the pharynx to the gastroesophageal junction. A lack of peristalsis in the esophagus can lead to hypertensive lower esophageal sphincter (LES). Surgical interventions to remedy long-gap esophageal atresia are often marred by dysmotility and impaired quality of life. In the case of bioengineered esophageal replacements, the restoration of physiological functionality must meet the requirements of both gravitational and peristaltic food transport. This becomes challenging due to the phenomenon of at-will “primary peristalsis,” a complex interplay between the central and enteric nervous systems.
Early reports in esophageal wall replacement demonstrated no muscular ingrowth with non-absorbable materials like polytetrafluoroethylene or Dacron. Surface functionalization of these bio-inert prosthetic materials with antigenic collagen typically result in a moderate cellular repopulation at best. Moreover, major side effects associated with the use of these materials have been reported, including stricture formation and inflammatory reactions.
Absorbable biomaterials have also been proposed as esophageal prosthetics to improve biocompatibility and minimize the host-inflammatory response exhibited with fluoropolymers. These were typically extra cellular matrix patches or collagen matrices derived from the urinary bladder or intestinal submucosa. The use of acellular xenogenic extra cellular matrix scaffolds to repair patch defects in the esophageal wall of canine models demonstrated neovascularization and neo-innervation, but no repopulation of esophageal smooth muscle.
Acellular approaches were improved by seeding biomaterials with cells. A modular approach to the regeneration of the esophagus by Saxena et al. used basement membrane matrix coated scaffolds to promote survival and unidirectional alignment of both epithelial cells as well as smooth muscle. Autologous neo-esophagus constructs have been engineered using composite cells (human esophageal epithelial cells, aortic smooth muscle cells and dermal fibroblasts) embedded into porcine tendon collagen or PGA meshes. More recently, Nakase et al. replaced a small portion of resected esophagus using keratinocytes, fibroblasts and smooth muscle cells seeded on human amniotic membrane and PGA sheets.
Although these attempts at tissue engineering displayed better repopulation of constituent cell types and similarities to native esophagus morphology, most segments remain aperistaltic and may cause dysmotility related problems during long-term implantation. In order to externally induce peristalsis, an artificial esophagus have been engineered using nickel-titanium shape memory alloys, and programmed to display peristaltic patterns when implanted in a goat model. Independent experiments using these materials for esophageal reconstruction, however, resulted in stenosis to different degrees. It appears that the paradigm of functional esophageal tissue engineering, if clinically intended to replace long segments, must mandatorily include peristalsis mediated by the intramural and myogenic esophageal components.
Similar problems have plagued attempts to reconstruct intestinal structures. The small intestine is the primary nutrient absorptive structure of the GI tract. Peristalsis and segmental contractions of the small intestine increase the surface area to facilitate greater absorption by the villi of the intestinal epithelium. Loss of intestinal segments due to congenital defects or multiple surgical resections due to inflammation or cancer result in short bowel syndrome. Short segments of small bowel result in malabsorption, malnutrition and adaptive alteration of motility patterns.
Tissue engineering also offers a possible advance to the bowel lengthening surgeries commonly carried out in short bowel syndrome. Collagen sponge scaffolds seeded with autologous smooth muscle cells have been successfully implanted as patch grafts in canine models. These patch grafts regenerated the mucosal and intestinal epithelial layers along with the villi structures. The major problem encountered with these grafts, however, was shrinkage. Dunn et al. used pseudo-tubular structures formed from collagen sponge scaffolds seeded with intestinal smooth muscle cells. The tubular structures were neovascularized within a month after prevascularization in the omentum. Unfortunately, these techniques did not regenerate the enteric neuronal layers, and the smooth muscle cells demonstrated a phenotypic switch to their non-contractile forms.
In one attempt to mimic the epithelium-mesenchyme interactions of GI tract structures, intestinal organoid units have been cultured and seeded onto tubular polymer scaffolds. Vacanti and colleagues implanted tissue-engineered intestine comprised of neonatal rat intestinal organoid units into the omentum of adult rats, and then subsequently implanted these constructs to rescue morbidity resulting from a massive bowel resection. Scaffolds made of small intestinal submucosa and wrapped with omentum were implanted in canine models of short bowel syndrome. These scaffolds repaired patch defects and replaced tubular segments of short bowel, thereby increasing the length of the short bowel. Tissue engineered small intestinal constructs regenerated enteric neuronal plexuses and met basic physiological demands. However, these techniques did not regenerate the alignment of the circular and longitudinal smooth muscle that is crucial to generating appropriate force and motility to facilitate nutrient absorption.
Regeneration of colon segments is similarly elusive. The colon is contiguous with the small intestine, facilitating water absorption and excretion of stool. Loss of colonic segments by surgical resections e.g., to treat aganglionosis (Hirschsprung's Disease) or inflammation significantly alters colonic motility. Disruption of colonic motility alters transit time, resulting in constipation or diarrhea. The idiopathic nature of some of these disease states poses a strong demand for in vitro tissue engineered models of colon, where investigations can be carried out on individual components (smooth muscle, enteric neurons, interstitial cells and mucosa) to understand alterations in pathophysiological conditions. Moreover, alterations in peristalsis and segmental contractions of the colon have direct implications on an individual's quality of life.
Recently, Vacanti et al. reported a tissue engineered colon construct using composite poly lactic and glycolic acid polymers seeded with organoid units isolated from the sigmoid colon. They demonstrated that the tissue engineered conduits have significant absorptive capacity when implanted into animals, but there was no direct measurement of peristalsis or motility.
Although significant advances have been made in tissue engineering of tubular structures, there is a need for better solutions in regeneration of functional smooth muscle structures to maintain various aspects of physiology, like peristalsis, contraction and relaxation. Accordingly, there also exists a need for better techniques for bioengineering of tubular tissues.